Microscopic Description of Protein-RNA Interactions in Nucleoprotein Condensates

Abstract

All eukaryotic cells use membrane-less compartments to spatially confine critical cellular processes, thereby increasing their throughput. A growing number of studies suggest that such compartmentalization occurs in response to cellular signals via a physical mechanism akin to liquid-liquid phase separation. Fused in sarcoma (FUS) is one of the best-studied proteins known to undergo a liquid-liquid phase separation, forming a biological condensate that recruits specific biomolecules implicated in DNA repair. Recent experiments have found the physical properties of such condensates to depend sensitively on the presence of nucleic acids in the condensate and on point mutations to the FUS sequence. A number of such point mutations have been implicated in neurodegenerative diseases including amyotrophic lateral sclerosis and dementia, for which there is no cure. By carrying out long time scale, all-atom molecular dynamics simulations of FUS protein systems, this study provides the first atomic-level insights into the structure of the biological condensate. The study's primary objective was to identify specific interactions within the FUS proteins, as well as between the proteins and RNA, that control the condensate's viscosity. Microsecond time scale simulations of a single FUS protein in solution and of an aggregate of two FUS proteins revealed the formation of persistent arginine-tyrosine contacts that were previously reported to orchestrate FUS aggregation. Uracil RNA homopolymers of varying lengths were then introduced within the FUS aggregate and simulated using the all-atom molecular dynamics method. The simulations have shown that binding of FUS to RNA depends on the length of the RNA, suggesting a pathway for RNA import into the condensate. Finally, we have applied the steered molecular dynamics protocol to pull bound RNA molecules out of the condensate, which has elucidated the critical molecular interactions holding the condensates together.